The aims of this project are to fractionate, isolate, and biochemically characterize allergenic components of Bermuda grass (Cynodon dactylon) pollen, and to use major and minor allergenic components of Bermuda for study of cross reactivity between Bermuda allergens and allergens in other grass pollens. We propose to confirm studies of Bermuda allergens by Orren and Dowdle, and extend them to fully characterize all chemically distinct components with allergenic properties. The extent to which heterogeneity results from aggregation sulfhydryl oxidation and deamination will be explored. Other aims include use of purified Bermuda allergens as reference materials in development of methods for standardization of Bermuda pollen extracts. A single large lot of Bermuda pollen will be employed for all studies. Fractionation techniques will include gel filtration chromatography, ion exchange chromatography, and preparative isoelectric focusing. At each fractionation step, specific activities of major fractions will be tested by skin prick testing by the end-point dilution method, on a panel of at least 10 volunteers, to identify major allergenic components. Other tests of biological activity will include leukocyte histamine release and immunoassays for IgE antibodies in serum. Purification of allergenic fractions will also be tested by use of a rabbit model of reaginic antibody formation. The purity and physicochemical characteristics of isolated allergens will be tested by several procedures including SDS polyacrylamide electrophoresis, isoelectric focusing in polyacrylamide gels, and by in vitro and in vivo immunological methods. Cross reactivity with pollen allergens from other grass species will be tested, employing immunoassay inhibition techniques and immunodiffusion in agar gel slides to study interaction between purified Bermuda components, monospecific antisera, and pollen extracts. These methods will identify allergens that are unique to Bermuda pollen that will be used to characterize and standardize crude Bermuda pollen extracts.